The binding of epidermal growth factor (EGF) to its plasma membrane receptor initiates a broad array of cellular responses, ultimately culminating in mitosis. The receptor is a transmembrane glycoprotein of M/r approximately 170,000, the sequence of which has been deduced from the cDNA for the receptor. A second natural ligand for the EGF receptor is transforming growth factor-alpha (TGFalpha), which has sequence and 3-dimensional structural homology with EGF. When EGF or TGFalpha binds to the extracytoplasmic domain of the receptor, a protein tyrosine kinase within the cytoplasmic domain of the receptor is activated, and the receptor dimerizes. Activation of the kinase on binding of EGF or TGFalpha to the receptor is generally considered to be an important step in signal transduction; however, the ultimate biological responses to EGF and TGFalpha, while similar, are not identical. The long term goal of this project is to develop an in-depth understanding of the protein chemistry and enzymology of the EGF receptor as it relates to the mechanism of signal transduction. This proposal addresses this long term goal through three specific aims: (1) To identify residues of the receptor adjacent to bound EGF or TGFalpha by a strategy of affinity cross-linking of site-directed mutants of EGF or TGFalpha to the receptor and determination of the sites of cross-linking by protein microsequencing, thereby testing whether the differences in biological response to EGF and TGFalpha correlate to subtle differences in binding sites. (2) To identify receptor residues important in EGF and/or TGFalpha recognition by a strategy of selective site-directed mutagenesis of the receptor and quantitative EGF and TGFalpha binding assays of mutant receptors. (3) To identify residues within the ATP binding site of the receptor responsible for chemical and thermal inactivation of receptor kinase activity by site-directed mutagenesis and testing the resulting mutant receptors for altered properties of chemical and thermal inactivation.